Staphylococcus aureus produces a number of toxins of great medical importance, including toxic shock syndrome toxin (TSST-1) and enterotoxins B and C. Several of these toxins are carried on so-called pathogenicity islands (SaPIs), genetic elements that resemble prophages, but lack most of the genes normally associated with phage production. These elements are normally stable, but can be mobilized by infection with certain phages or by the induction of indigenous prophages, upon which the SaPIs get excised and packaged into phage-like particles that can transfer the toxin genes to new cells. The type member of the SaPI family, SaPI1 can be mobilized by phage 80? and is efficiently packaged into phage-like particles using 80? structural proteins. However, the SaPI1 capsid is only 1/3 the size of the normal 80? capsid, commensurate with its smaller genome. With the emergence of antibiotic-resistant Staphylococcus strains and the renewed interest in therapeutic use of bacteriophages to treat such infections, the phage-induced mobilization of SaPI1 will be an important consideration. The overall objective of this study is to understand the structural basis for the phage-induced mobilization and spread of S. aureus pathogenicity islands and its role in the development of bacterial pathogenesis. In this project, we address two specific questions in relation to the SaPI1 mobilization: How does SaPI1 induce the formation of small capsids from 80?-encoded proteins? How is the SaPI1 DNA selected for packaging into these capsids? Understanding these questions will have a bearing on understanding the evolution of pathogenicity in S. aureus. To this end, we will use a combination of genetic, biochemical and structural approaches, including cryo-EM, X-ray crystallography and Mass Spectrometry. The specific aims of this study are: (1) Define the role of the 80? structural gene products in capsid formation (2) Determine the mechanism of SaPI1-induced capsid size determination (3) Identify the determinants for DNA discrimination by SaPI1. PUBLIC HEALTH RELEVANCE: Staphylococcus aureus has become a major health problem in hospitals, especially with the emergence of multiple antibiotic resistant strains. Pathogenic S. aureus may cause severe systemic infections by producing several toxins from genes carried on so-called pathogenicity islands in the bacterial genome. The bacteriophage/pathogenicity island system under study in this project is of special interest for its role in the horizontal spread and long-term establishment of pathogenicity in the population.